Extreme Thermostable T4 Polynucleotide Kinase (ET T4 PNK)
$285.00 - $1,085.00
All products have special prices for bulk purchase, please contact for more details if required.
Cat. No.: ETT4PNK-500 (for 500U)
Cat. No.: ETT4PNK-2500 (for 2500U)
Description
Extreme Thermostable T4 Polynucleotide Kinase (ET T4 PNK) is our proprietary, innovative enzyme—a thermostable homolog of T4 PNK. It retains the same function as T4 PNK, catalyzing the transfer of the γ-phosphate group from ATP to the 5’-hydroxyl end of oligonucleotides, including double-stranded or single-stranded DNA and RNA.
This enzyme exhibits exceptional heat stability, maintaining full activity even after 30 minutes at 95°C. It operates within a temperature range of 50–95°C, though its activity significantly decreases below 65°C. Within 75–95°C, it maintains consistent enzymatic activity. These unique properties enable efficient processing of nucleic acid substrates with highly structured and complex configurations, as well as experimental conditions requiring extreme temperatures.
ET T4 PNK possesses 5’-kinase activity but lacks 3’-kinase and phosphatase activity.
Applications
Phosphorylation of DNA or RNA with highly complex secondary structures at the 5’ terminus.
One-step high-temperature phosphorylation and ligation of DNA or RNA.
Activity Definition
One unit of ET T4 PNK is defined as the amount of enzyme required to catalyze the phosphorylation of 20 pmol of oligo DNA within 30 minutes at 75°C.
Usage Instructions
Reaction Setup:
| Component | Amount |
|---|---|
DNA (5’-end) | 5–50 pmol |
10× PNKnase Buffer | 2.5 μl |
10 mM ATP | 1 μl |
ET T4 PNK(10 U/μl) | 1 μl |
ddH₂O | Up to 25 μl |
Incubation: Incubate at 75°C for 15–30 minutes.
Precautions
- DNA Quantification: 1 pmol of 5’-end DNA corresponds to 1 µg of a 3 kb PCR product.
- Evaporation Control: Due to the enzyme’s high working temperature, it is recommended to use a PCR instrument with a heated lid to minimize sample evaporation.
- Salt Sensitivity: ET PNKnase is sensitive to salt concentrations. When Na⁺ or K⁺ levels exceed 100 mM, enzymatic activity declines sharply. The recommended range is 0–50 mM.
- Enzyme Inactivation: To inactivate the enzyme, digest with 0.2 mg/ml of proteinase K.
Storage
Store at -20°C for up to 3 years.
Related:
- T4 Polynucleotide Kinase Kit
- T4 Polynucleotide Kinase
- T4 Polynucleotide Kinase (3' phosphatase minus)
- Extreme Thermostable T4 Polynucleotide Kinase (ET T4 PNK)
